The renal distal nephoron plays an important role in the regulation of systemic acid-base balance. The tubule segments that comprise this portion of the nephron reabsorb the fraction of filtered bicarbonate not reclaimed by the proximal tubule and further acidify the urine such that net urinary acid excretion matches the daily acid load produced by metabolism. Although much is known about the transepithelial movement of hydrogen and bicarbonate across the distal nephron, the cellular mechanism(s) of acid transport remain largely unknown. It is the purpose of this proposal to examine directly the cellular mechanisms udnerlying H+ and HCO3-transport by the distal tubule. To this end electrophysiological techniques will be employed on isolated and perfused late distal tubules of Amphiuma in vitro. In particular, studies will be conducted with voltage recording and ion-sensitive microelectrodes. The late distal tubule of Amphiuma is ideally suited for this study due to the high rate of net H+ secretion and large homogenous cells which are amenable to impalements with microelectrodes The specific aims of this proposal include: (1) measuring transepithelial net fluxes of H+ so as to define the transport properties of this epithelium and (2) defining the mechanism(s) of H+ and HC03-movement across the luminal and basolateral membrane. This will involve measurements of the recovery of intracellular pH subsequent to intracellular acid loading. In addition, other intracellular electrolytes (e.g. Na+, C1-) will be monitored to examine the coupling of H+ and HC03-transport to these ions. The studies will provide insight into the cellular mechanisms of net H+ and HC03-transport in the distal nephron, and provide a framework for future studies on the regulation of this process. Moreover, information on the general phenomenon of intracellular pH regulation will be obtained which should be of interest beyond the scope of renal physiology.